关键词: 谱法, 高压液相, 荧光计, 苯并芘, 胚胎, 哺乳动物, 妊娠初期

Abstract: Objective：To establish a high performance liquid chromatography with fluorescence detection (HPLC-FLD) for the detection of Benzo(a)pyrene in the embryonic tissues of rat early pregnancy, so as to provide experimental data for future clinical application. Methods：The level of Benzo(a)pyrene in the embryonic tissues of rats was determined by Shimadzu HPLC with Shimadzu RF-535 fluorescence detector. The samples were extracted by acetonitrile, without cleaned-up. The chromatographic separation was performed using a column of Kromasil C18 (250 mm×4.6 mm, 5 μL) with a mobile phase consisting of methanol and water (95∶5, V/V) at the flow rate of 1.0 mL/min. Excitation wavelength (Ex) of fluorescence detection was 364 nm, the emission wavelength (Em) 427 nm, and the injection volume 20 μL. Results：The level of Benzo(a)pyrene in the tissues of rats was linear in the range of 0.002 μg/mL to 0.5 μg/mL. The regression equation of standard curve of extraction was ■=426 174x－316.95 with a correlation coefficient of 0.999 9. The average recovery for Benzo(a)pyrene standard spiked was more than 95% while the inter-day precision RSD was 3.38%, the intra-day precision RSD was 2.80%, and the stability (n=6) was 2.58%. The benzo(a)pyrene concentration of samples was in the range of 0-0.5837 μg/g. Conclusions：The HPLC-FLD method has the advantages of simple operation, low experiment cost, and coincident veracityand sensitivity. The process of sample extraction is simple and time-saving. We conclude that the HPLC-FLD may be a suitable way for the detection of benzo(a)pyrene in samples such as embryonic tissues in the clinical practice and research projects.

Key words: Chromatography, high pressure liquid, Fluorometry, Benzo(a)pyrene, Embryo, mammalian, Pregnancy trimester, first